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The article deals with the first record of a humpback whale stranding in October 2008 in the northern coast of Brazil. It cites the occurrence of such whales primarily in coastal and continental shelf waters. The area where the whale was found stranding is said to comprise of a coast of indentation, formed by lowlands with several indentations. The carcass of the whale has a measurement of 10 meters in length and was in advanced state of decomposition.

The composite material-like extracellular matrix (ECM) in the sinoatrial node (SAN) supports the native pacemaking cardiomyocytes (PCMs). To test the roles of SAN ECM in the PCM phenotype and function, we engineered reconstructed-SAN heart tissues (rSANHTs) by recellularizing porcine SAN ECMs with hiPSC-derived PCMs. The hiPSC-PCMs in rSANHTs self-organized into clusters resembling the native SAN and displayed higher expression of pacemaker-specific genes and a faster automaticity compared with PCMs in reconstructed-left ventricular heart tissues (rLVHTs). To test the protective nature of SAN ECMs under strain, rSANHTs and rLVHTs were transplanted onto the murine thoracic diaphragm to undergo constant cyclic strain. All strained-rSANHTs preserved automaticity, whereas 66% of strained-rLVHTs lost their automaticity. In contrast to the strained-rLVHTs, PCMs in strained-rSANHTs maintained high expression of key pacemaker genes (HCN4, TBX3, and TBX18). These findings highlight the promotive and protective roles of the composite SAN ECM and provide valuable insights for pacemaking tissue engineering., Competing Interests: Declaration of interests D.K.L. is a consultant for Novoheart, Ltd., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Directed cardiomyogenesis from human induced pluripotent stem cells (hiPSCs) has been greatly improved in the last decade but directed differentiation to pacemaking cardiomyocytes (CMs) remains incompletely understood. In this study, we demonstrated that inhibition of NODAL signaling by a specific NODAL inhibitor (SB431542) in the cardiac mesoderm differentiation stage downregulated PITX2c, a transcription factor that is known to inhibit the formation of the sinoatrial node in the left atrium during cardiac development. The resulting hiPSC-CMs were smaller in cell size, expressed higher pro-pacemaking transcription factors, TBX3 and TBX18, and exhibited pacemaking-like electrophysiological characteristics compared to control hiPSC-CMs differentiated from established Wnt-based protocol. The pacemaker-like subtype increased up to 2.4-fold in hiPSC-CMs differentiated with the addition of SB431542 relative to the control. Hence, Nodal inhibition in the cardiac mesoderm stage promoted pacemaker-like CM differentiation from hiPSCs. Improving the yield of human pacemaker-like CMs is a critical first step in the development of functional human cell-based biopacemakers., (Published by Elsevier B.V.)

Genetically encoded fluorescent voltage indicators, such as ArcLight, have been used to report action potentials (APs) in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). However, the ArcLight expression, in all cases, relied on a high number of lentiviral vector-mediated random genome integrations (8-12 copy/cell), raising concerns such as gene disruption and alteration of global and local gene expression, as well as loss or silencing of reporter genes after differentiation. Here, we report the use of clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 nuclease technique to develop a hiPSC line stably expressing ArcLight from the AAVS1 safe harbor locus. The hiPSC line retained proliferative ability with a growth rate similar to its parental strain. Optical recording with conventional epifluorescence microscopy allowed the detection of APs as early as 21 days postdifferentiation, and could be repeatedly monitored for at least 5 months. Moreover, quantification and analysis of the APs of ArcLight-CMs identified two distinctive subtypes: a group with high frequency of spontaneous APs of small amplitudes that were pacemaker-like CMs and a group with low frequency of automaticity and large amplitudes that resembled the working CMs. Compared with FluoVolt voltage-sensitive dye, although dimmer, the ArcLight reporter exhibited better optical performance in terms of phototoxicity and photostability with comparable sensitivities and signal-to-noise ratios. The hiPSC line with targeted ArcLight engineering design represents a useful tool for studying cardiac development or hiPSC-derived cardiac disease models and drug testing., (©AlphaMed Press 2019.)

Objectives: Several neurotransmitters involved in brain development are altered in fragile X syndrome (FXS), the most common monogenic cause of autism spectrum disorder (ASD). Serotonin plays a vital role in synaptogenesis and postnatal brain development. Deficits in serotonin synthesis and abnormal neurogenesis were shown in young children with autism, suggesting that treating within the first years of life with a selective serotonin reuptake inhibitor might be the most effective time. In this study we aimed to identify molecular biomarkers involved in the serotonergic pathway that could predict the response to sertraline treatment in young children with FXS., Methods: Genotypes were determined for several genes involved in serotonergic pathway in 51 children with FXS, ages 24-72months. Correlations between genotypes and deviations from baseline in primary and secondary outcome measures were modeled using linear regression models., Results: A significant association was observed between a BDNF polymorphism and improvements for several clinical measures, including the Clinical Global Impression scale (P=0.008) and the cognitive T score (P=0.017) in those treated with sertraline compared to those in the placebo group. Additionally, polymorphisms in the MAOA, Cytochrome P450 2C19 and 2D6, and in the 5-HTTLPR gene showed a significant correlation with some of the secondary measures included in this study., Conclusion: This study shows that polymorphisms of genes involved in the serotonergic pathway could play a potential role in predicting response to sertraline treatment in young children with FXS. Larger studies are warranted to confirm these initial findings., (Copyright © 2017 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Fragile X syndrome (FXS) affects individuals with more than 200 CGG repeats (full mutation) in the fragile X mental retardation 1 (FMR1) gene. Those born with FXS experience cognitive and social impairments, developmental delays, and some features of autism spectrum disorders. Carriers of a premutation (55-200 CGG repeats) are generally not severely affected early in life; however, are at high risk of developing the late onset neurodegenerative disorder, Fragile X-associated Tremor/Ataxia Syndrome (FXTAS), or Fragile X-associated Primary Ovarian Insufficiency (FXPOI), and may have other medical conditions such as developmental delay, autism spectrum disorders, hypertension, anxiety, and immune-mediated disorders. Here we present a case of a 58-year-old man with a borderline IQ, average memory skills, and executive function deficits. He met criteria for multiple psychiatric diagnoses and presented with tremor and ataxia, meeting criteria for FXTAS. Molecular testing unveiled a completely unmethylated FMR1 full mutation in peripheral blood mononucleated cells with elevated FMR1 mRNA and premutation alleles of different sizes in two other tissues (primary fibroblasts and sperm), indicating the presence of allele instability based on both inter- and intra-tissue mosaicism. The observation of FXTAS in this case of a full mutation mosaic man suggests that the pathogenic mechanism underlying this disorder is not observed exclusively in premutation carriers as it was originally thought. The concomitant presence of features of FXS and late onset neurological deterioration with probable FXTAS likely result from a combined molecular pathology of elevated FMR1 mRNA levels, a molecular hallmark of FXTAS and low FMRP expression that leads to FXS., (© 2015 Wiley Periodicals, Inc.)

Background: The diagnosis of 22q11 deletion syndrome (22q11DS) is often delayed or missed due to the wide spectrum of clinical involvement ranging from mild to severe, often life-threatening conditions. A delayed diagnosis can lead to life-long health issues that could be ameliorated with early intervention and treatment. Owing to the high impact of 22q11DS on public health, propositions have been made to include 22q11DS in newborn screening panels; however, the method of choice for detecting 22q11DS, fluorescent in situ hybridization, requires specialized equipment and is cumbersome for most laboratories to implement as part of their routine screening. We sought to develop a new genetic screen for 22q11DS that is rapid, cost-effective, and easily used by laboratories currently performing newborn screening., Methods: We evaluated the accuracy of multiplex droplet digital PCR (ddPCR) in the detection of copy number of 22q11DS by screening samples from 26 patients with 22q11DS blindly intermixed with 1096 blood spot cards from the general population (total n = 1122)., Results: Multiplex ddPCR correctly identified all 22q11DS samples and distinguished between 1.5- and 3-Mb deletions, suggesting the approach is sensitive and specific for the detection of 22q11DS., Conclusions: These data demonstrate the utility of multiplex ddPCR for large-scale population-based studies that screen for 22q11DS. The use of samples from blood spot cards suggests that this approach has promise for newborn screening of 22q11DS, and potentially for other microdeletion syndromes, for which early detection can positively impact clinical outcome for those affected., (© 2014 American Association for Clinical Chemistry.)

Expansions of more than 200 CGG repeats (full mutation) in the FMR1 gene give rise to fragile X syndrome (FXS) through a process that generally involves hypermethylation of the FMR1 promoter region and gene silencing, resulting in absence of expression of the encoded protein, FMRP. However, mosaicism with alleles differing in size and extent of methylation often exist within or between tissues of individuals with FXS. In the current work, CGG-repeat lengths and methylation status were assessed for eighteen individuals with FXS, including 13 mosaics, for which peripheral blood cells (PBMCs) and primary fibroblast cells were available. Our results show that for both PBMCs and fibroblasts, FMR1 mRNA and FMRP expression are directly correlated with the percent of methylation of the FMR1 allele. In addition, Full Scale IQ scores were inversely correlated with the percent methylation and positively correlated with higher FMRP expression. These latter results point toward a positive impact on cognition for full mutation mosaics with lower methylation compared to individuals with fully methylated, full mutation alleles. However, we did not observe a significant reduction in the number of seizures, nor in the severity of hyperactivity or autism spectrum disorder, among individuals with mosaic genotypes in the presentation of FXS. These observations suggest that low, but non-zero expression of FMRP may be sufficient to positively impact cognitive function in individuals with FXS, with methylation mosaicism (lowered methylation fraction) contributing to a more positive clinical outcome.

Carriers of the fragile X premutation (FPM) have CGG trinucleotide repeat expansions of between 55 and 200 in the 5'-UTR of FMR1, compared to a CGG repeat length of between 5 and 54 for the general population. Carriers were once thought to be without symptoms, but it is now recognized that they can develop a variety of early neurological symptoms as well as being at risk for developing the late onset neurodegenerative disorder fragile X-associated tremor/ataxia syndrome (FXTAS). Several mouse models have contributed to our understanding of FPM and FXTAS, and findings from studies using these models are summarized here. This review also discusses how this information is improving our understanding of the molecular and cellular abnormalities that contribute to neurobehavioral features seen in some FPM carriers and in patients with FXTAS. Mouse models show much of the pathology seen in FPM carriers and in individuals with FXTAS, including the presence of elevated levels of Fmr1 mRNA, decreased levels of fragile X mental retardation protein, and ubiquitin-positive intranuclear inclusions. Abnormalities in dendritic spine morphology in several brain regions are associated with neurocognitive deficits in spatial and temporal memory processes, impaired motor performance, and altered anxiety. In vitro studies have identified altered dendritic and synaptic architecture associated with abnormal Ca(2+) dynamics and electrical network activity. FPM mice have been particularly useful in understanding the roles of Fmr1 mRNA, fragile X mental retardation protein, and translation of a potentially toxic polyglycine peptide in pathology. Finally, the potential for using these and emerging mouse models for preclinical development of therapies to improve neurological function in FXTAS is considered.

The aim of this study was to assess the effect of chemical composition, coagulation properties, pH, and titratable acidity (TA, SH°/50 ml) of vat milk on Grana Padano cheese yield (CY) under field conditions. Twelve cheese-making sessions were carried out from February to December 2009 in a dairy cooperative of Grana Padano Consortium (Italy), for a total of 96 vats of milk processed. For each vat, samples of raw milk were collected and analysed for quality traits (fat, protein, and casein contents), pH, TA, and milk coagulation properties (MCP), measured as rennet coagulation time (RCT, min), curd-firming time (k(20), min), and curd firmness (a(30), mm). Cheese yield was expressed as kilograms of cheese per 100 kg milk transformed, and was measured after 2 d of drainage. Fat, protein, and casein contents were positively and strongly correlated with CY (coefficients of correlation, r = 0.72, 0.88, and 0.84, respectively; P < 0.001). Coagulation properties were moderately and significantly (P < 0.001) related to CY: milk that coagulated earlier and had stronger a(30) was associated to greater CY. Cheese yield was analysed with a model that accounted for fixed effects of cheese-making day, fat and protein content, TA, and a(30). Significance was found for all the effects (P < 0.05). Milk characterised by high values of a(30) resulted in higher CY than milk with low values of a 30, indicating that MCP could be used as indicators of cheese-making efficiency. Future research should investigate the relationships between MCP and quality of cheese, and explore the feasibility of including MCP in multiple component milk pricing system for Grana Padano cheese production.

Loss of the RecQ DNA helicase WRN protein causes Werner syndrome, in which patients exhibit features of premature aging and increased cancer. WRN deficiency induces cellular defects in DNA replication, mitotic homologous recombination (HR), and telomere stability. In addition to DNA unwinding activity, WRN also possesses exonuclease, strand annealing, and branch migration activities. The single strand binding proteins replication protein A (RPA) and telomere-specific POT1 specifically stimulate WRN DNA unwinding activity. To determine whether RPA and POT1 also modulate WRN branch migration activity, we examined biologically relevant mobile D-loops that mimic structures in HR strand invasion and at telomere ends. Both RPA and POT1 block WRN exonuclease digestion of the invading strand by loading on the strand. However, only RPA robustly stimulates WRN branch migration activity and increases the percentage of D-loops that are disrupted. Our results are consistent with cellular data that support RPA enhancement of branch migration during HR repair and, conversely, POT1 limitation of inappropriate recombination and branch migration at telomeric ends. This is, to our knowledge, the first evidence that RPA can stimulate branch migration activity.

Objective: To evaluate the relationship between PD and ACS and the association of PD and periodontitis in ACS patients., Methods: Fifty-eight ACS patients and 57 controls with no history of coronary artery disease (CAD) were included in the study., Variables: arterial hypertension, diabetes, dyslipidemia, obesity, history of CAD, cigarette smoking, and interleukin-1beta gene polymorphism., Results: One hundred and fifteen subjects were enrolled in the study. In the ACS group, 58 patients were evaluated, 32 of whom (55.2%) were male and 26 (44.8%), female. In the control group, 57 subjects were evaluated, 32 (56.1%) of whom were male and 25 (43.9%), female. Periodontal disease was diagnosed in 26 (44.8%) ACS patients and 15 (26.6%) control patients (beta2 = 4.43, p = 0.04). In a logistic regression analysis, the odds ratio for association between PD and ACS was 1.8 (95% CI: 1.0-5.0); p = 0.24. The odds ratio for association of periodontitis with ACS was 4.5 (95% CI: 1.3-15.6); p = 0.019., Conclusion: No independent association was found between PD and ACS. There was an independent association between periodontitis and ACS.

Usher syndrome type IIA (MIM: 27601) is an autosomal recessive disorder characterized by moderate to severe congenital deafness and progressive retinitis pigmentosa. We recently identified the human Usher syndrome type IIA gene (USH2A) on chromosome 1q41, which encodes a protein possessing 10 laminin epidermal growth factor and four fibronectin type 3 domains, both commonly observed in extracellular matrix proteins. To gain insight into the pathogenesis of Usher syndrome type IIA, we isolated and characterized the murine (Ush2a) and rat (rat Ush2a) orthologs of human USH2A. We mapped mouse Ush2a by fluorescence in situ hybridization to mouse chromosome 1 in the region syntenic to human chromosome 1q41. Rat Ush2a has been localized by radiation hybrid mapping to rat chromosome 13 between d13rat49 and d13rat76. The mouse and rat genes, similar to human USH2A, are expressed primarily in retina and cochlea. Mouse Ush2a encodes a 161-kDa protein that shows 68% identity and 9% similarity to the human USH2A protein. Rat Ush2a encodes a 167-kDa protein with 64% identity and 10% similarity to the human protein and 81% identity and 5% similarity to the mouse USH2A protein. The predicted amino acid sequence of the mouse and rat proteins, like their human counterpart, contains a leader sequence, an amino-terminal globular domain, 10 laminin epidermal growth factor domains, and four carboxy-terminal fibronectin type III motifs. With in situ hybridization, we compared the cellular expression of the USH2A gene in rat, mouse, and human retinas. USH2A mRNA in the adult rat, mouse, and human is expressed in the cells of the outer nuclear layer of the retina, one of the target tissues of the disease. In the developing rat retina, Ush2a mRNA expression appears in the neuroepithelium at embryonic day 17.